Transplanted tissues or organs are always rejected by recipients unless they take immunosuppressive drugs for the rest of their lives. However, traditional immunosuppressive drugs cause serious side-effects including viral infection and malignancy. Achieving long-term allograft survival or tolerance in the absence of continuous global immunosuppression is a highly desirable goal in transplantation. ESAT-6 is a protein of 6 kDa that was originally identified as a T cell Ag in the short-term culture filtrate of M. tuberculosis. Substantial evidence has shown that it is a virulence factor that promotes the growth and pathogenesis of M. tuberculosis. In particular, ESAT-6 has been shown to suppress T cell proliferation. Our preliminary studies for the first time have demonstrated that ESAT-6 promotes the induction of long-term islet allograft survival and suppresses graft-infiltrating T cell proliferation. Therefore, ESAT-6 could be utilized as a novel agent to induce long- term allograft survival or tolerance in the absence of continuous treatment with a globally immunosuppressive agent. Based on these findings, we propose to further study the mechanisms underlying the suppression of allograft rejection by ESAT-6 with two specific aims: (1) To investigate the mechanisms by which ESAT-6 suppresses alloimmune responses in vivo and in vitro. We will investigate the impact of ESAT-6 on T helper cell differentiation into Th1/Th17/Th2 cells and on T cell development into regulatory T (Treg) and memory T (Tmem) cells in the context of alloimmune responses, as tipping their balance toward either Th2 or Treg regulation is critical for long-term allograft survival; and (2) To study whether ESAT-6 also promotes long-term islet allograft survival in diabetic NOD mice in the absence or presence of brief and mild immunosuppression. The goal of this proposal is to explore new strategies to induce long-term allograft survival or tolerance in the absence or presence of brief and mild immunosuppression using a novel protein that is originally produced by Mycobacteria tuberculosis to enhance their growth. This study will look into two basic sets of immunologic balance: Th1/Th17/Th2 and Treg/Tmem, both of which are critical for determining an allograft fate. Therefore this project has important clinical implications for the cure of type 1 diabetes by islet transplantation and perhaps other end-stage organ diseases via solid organ transplantation.